The structure and function of heme proteins has been an area of intense investigation for decades because of their involvement in the most fundamental biological processes such as oxygen transport, oxidative phosphorylation and photosynthesis. A large number of cytochromes, one-electron carrying heme proteins, are known that are involved in electron transport chains of all that the most anaerobic of organisms. Among these proteins, the cytochromes C constitute a group having very similar structure, with a few exceptions, but very different properties. It is a long range goal of this laboratory to determine how the structure of different cytochromes allow them to function optimally in their respective systems. The specificity involves both the matching of redox potentials to those of the other electron carriers in the system and the specificity of the binding interactions between them. Another interest of this laboratory is to develop 15N-NMR as a tool for the study of small proteins. Cytochromes C are appropriately sized proteins in which nitrogen atoms play unusually important roles in the structure and surface charge distribution of the protein. 15N can be incorporated biosynthetically into the cytochrome c2 of Rhodospirillum rubrum by supplying the organism with 15NH4 ion. The uniformly enriched cytochrome will be purified by existing procedures and its N-NMR spectrum examined as a function of pH, temperature and oxidation state. Resonance will be assigned as specifically as possible using known shifts of amino acid groups and double resonance techniques. Changes in chemical shifts of 15N resonances with pH will be correlated with ionizations known to occur in the molecule that are detected by changes in redox potential and visible absorption spectrum to determine which groups are capable of altering these properties. The time scale of the motions of resolved 15N resonances can be estimated from the value of the 1H-15N nuclear Overhouser effect. This information will be used to determine the rigidity of the different structural elements of the molecule. The local motion of the surface lysine residues might be used in future experiments to map out the site of interaction between the cytochrome and its redox partners.